Abstract
Fluorescence-based biosensors for membrane voltage (mV) allow dynamic optical recording of neuronal activity. Interestingly, the development of genetically encoded voltage indicators constitute a good alternative to classical voltage-sensitive dyes, thus allowing overcoming some of the inherent problems (e.g., optical noise, etc.) associated with these organic compounds. Here, we show the use of a genetically encoded voltage-sensitive fluorescent protein (VSFP), namely the VSFP2.32, which contains a mCerulean and Citrine tandem engaging in a constitutive fluorescent resonance energy transfer (FRET) process. By expressing VSFP2.32 in hippocampal cultured neurons, we were able to monitor mV alterations in single neurons by recording VSFP2.32 conformation-mediated FRET changes in a real-time mode.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Mutoh H, Knöpfel T (2013) Probing neuronal activities with genetically encoded optical indicators: from a historical to a forward-looking perspective. Pflugers Arch 465:361–371. doi:10.1007/s00424-012-1202-z
Borisov SM, Wolfbeis OS (2008) Optical biosensors. Chem Rev 108:423–461. doi:10.1021/cr068105t
Grinvald A, Hildesheim R (2004) VSDI: a new era in functional imaging of cortical dynamics. Nat Rev Neurosci 5:874–885. doi:10.1038/nrn1536
Knöpfel T, Díez-García J, Akemann W (2006) Optical probing of neuronal circuit dynamics: genetically encoded versus classical fluorescent sensors. Trends Neurosci 29:160–166. doi:10.1016/j.tins.2006.01.004
Day RN, Davidson MW (2009) The fluorescent protein palette: tools for cellular imaging. Chem Soc Rev 38:2887–2921. doi:10.1039/b901966a
Knöpfel T (2012) Genetically encoded optical indicators for the analysis of neuronal circuits. Nat Rev Neurosci 13:687–700. doi:10.1038/nrn3293
Mutoh H, Akemann W, Knöpfel T (2012) Genetically engineered fluorescent voltage reporters. ACS Chem Neurosci 3:585–592. doi:10.1021/cn300041b
Murata Y, Iwasaki H, Sasaki M et al (2005) Phosphoinositide phosphatase activity coupled to an intrinsic voltage sensor. Nature 435:1239–1243. doi:10.1038/nature03650
Perron A, Akemann W, Mutoh H, Knöpfel T (2012) Genetically encoded probes for optical imaging of brain electrical activity. Prog Brain Res 196:63–77. doi:10.1016/B978-0-444-59426-6.00004-5
Lundby A, Mutoh H, Dimitrov D et al (2008) Engineering of a genetically encodable fluorescent voltage sensor exploiting fast Ci-VSP voltage-sensing movements. PLoS One 3:e2514. doi:10.1371/journal.pone.0002514
Akemann W, Mutoh H, Perron A et al (2010) Imaging brain electric signals with genetically targeted voltage-sensitive fluorescent proteins. Nat Methods 7:643–649. doi:10.1038/nmeth.1479
Clark JD, Gebhart GF, Gonder JC et al (1997) Special report: the 1996 guide for the care and use of laboratory animals. ILAR J 38:41–48
Acknowledgements
This work was supported by Ministerio de Economía y Competitividad/ Instituto de Salud Carlos III (SAF2014-55700-P, PCIN-2013-019-C03-03 and PIE14/00034), Institució Catalana de Recerca i Estudis Avançats (ICREA Academia 2010), and Agentschap voor Innovatie door Wetenschap en Technologie (SBO-140028) to FC. Also, F.C., X.M. and V.F.-D. belong to the “Neuropharmacology and Pain” accredited research group (Generalitat de Catalunya, 2014 SGR 1251). We thank E. Castaño and B. Torrejón from the Scientific and Technical Services (SCT) group at the Bellvitge Campus of the University of Barcelona for their technical assistance.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Fernández-Dueñas, V., Morató, X., Knöpfel, T., Ciruela, F. (2016). Dynamic Recording of Membrane Potential from Hippocampal Neurons by Using a FRET-Based Voltage Biosensor. In: Luján, R., Ciruela, F. (eds) Receptor and Ion Channel Detection in the Brain. Neuromethods, vol 110. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3064-7_27
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3064-7_27
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3063-0
Online ISBN: 978-1-4939-3064-7
eBook Packages: Springer Protocols